title: A pathogenic nematode targets recognition proteins to avoid insect defenses.
abstract: Steinernemacarpocapsae is a nematode pathogenic in a wide variety of insect species. The great pathogenicity of this nematode has been ascribed to its ability to overcome the host immune response; however, little is known about the mechanisms involved in this process. The analysis of an expressed sequence tags (EST) library in the nematode during the infective phase was performed and a highly abundant contig homologous to serine protease inhibitors was identified. In this work, we show that this contig is part of a 641-bp cDNA that encodes a BPTI-Kunitz family inhibitor (Sc-KU-4), which is up-regulated in the parasite during invasion and installation. Recombinant Sc-KU-4 protein was produced in Escherichia coli and shown to inhibit chymotrypsin and elastase activities in a dose-dependent manner by a competitive mechanism with Ki values of 1.8 nM and 2.6 nM, respectively. Sc-KU-4 also inhibited trypsin and thrombin activities to a lesser extent. Studies of the mode of action of Sc-KU-4 and its effects on insect defenses suggest that although Sc-KU-4 did not inhibit the activation of hemocytes or the formation of clotting fibers, it did inhibit hemocyte aggregation and the entrapment of foreign particles by fibers. Moreover, Sc-KU-4 avoided encapsulation and the deposition of clotting materials, which usually occurs in response to foreign particles. We show by protein-protein interaction that Sc-KU-4 targets recognition proteins of insect immune system such as masquerade-like and serine protease-like homologs. The interaction of Sc-KU-4 with these proteins explains the ability of the nematode to overcome host reactions and its large pathogenic spectrum, once these immune proteins are well conserved in insects. The discovery of this inhibitor targeting insect recognition proteins opens new avenues for the development of S. carpocapsae as a biological control agent and provides a new tool to study host-pathogen interactions.
authors.affiliation: IBB/CBA and CIRN/Departamento de Biologia, Universidade dos Açores, Ponta Delgada, Portugal.
authors.forename: Duarte
authors.initials: D
authors.lastname: Toubarro
authors.forename: Mónica Martinez
authors.initials: MM
authors.lastname: Avila
authors.forename: Rafael
authors.initials: R
authors.lastname: Montiel
authors.forename: Nelson
authors.initials: N
authors.lastname: Simões
mesh_terms: D000818:Animals; D007611:Aprotinin / Q000235:genetics / Q000378:metabolism*; D015153:Blotting, Western; D020451:Contig Mapping; D017931:DNA Primers / Q000235:genetics; D004926:Escherichia coli; D019143:Evolution, Molecular; D020224:Expressed Sequence Tags / Q000378:metabolism*; D020869:Gene Expression Profiling; D057131:Immune Evasion / Q000235:genetics / Q000502:physiology*; D008958:Models, Molecular; D009348:Nematoda / Q000472:pathogenicity*; D011487:Protein Conformation; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization
chemical_list: D017931:DNA Primers; D007611:Aprotinin